In Amphiuma red blood cells, the Na/H and K/H antiports mediate volume regulation after shrinkage and swelling, respectively. These ionic flux routes play a crucial role in viability of certain tumor cells and are directly responsible for cell injury in cardiomyocites during hypoxic episodes. However, the basis for their activation and inactivation remains obscure. We hypothesize that antiport protein phosphorylation status determines activity, while phosphorylation pattern dictates ion selectivity. To test this hypothesis we will monitor transporter activity in conjunction with biochemical studies designed to evaluate transporter phosphorylation state in order to address the following specific aims: 1) Determine the effect of the phosphatase inhibitor Calyculin A in the ionic fluxes through the Na/H and K/H exchange. 2) Perform experiments to determine the time course for activation and inactivation of the Alkali Metal/H exchanger during hyperosmotic and hyposmotic perturbations and fit the data to a minimal two state model (active and inactive) proposed by Jennings, M.L. and Al-Rohil, N. (1990). 3) Determine which antiporter amino acid residues are phosphorylated or dephosphorylated in response to shrinkage or swelling.